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Zhou Y., Li H., Zhang P.-P., Wang L., Li Z.-F. (2018): Molecular mapping of leaf rust resistance gene LrL224 in Chinese wheat cultivar L224-3. Czech J. Genet. Plant Breed., 54: 65−70. Leaf rust, caused by Puccinia triticina, is a major wheat disease worldwide. The chinese wheat cultivar L224-3 showed high resistance to most of P. triticina pathotypes in the seedling and adult stage. Identifying and mapping the leaf rust resistance gene(s) in L224-3 is very useful for breeding leaf rust resistant wheat cultivars. In the present study, the wheat cultivar L224-3 and thirty-six lines with known leaf rust resistance genes were inoculated with 15 pathotypes at the seedling stage for gene postulation. A total of 144 F2:3 lines from the cross L224-3 × Zhengzhou 5389 were inoculated with the pathotype FHBQ for leaf rust genetic analysis at the seedling stage. A total of 1276 SSR (simple sequence repeat) markers and the STS (sequence tagged-site) marker ω-secali/Glu-B3 were used to test the parents, resistant and susceptible bulks. The polymorphic markers were used to genotype the F2:3 populations. L224-3 was highly resistant to all Lr26 avirulent pathotypes, showing the presence of Lr26 in L224-3. The presence of Lr26 in L224-3 was also confirmed by the molecular marker ω-secalin/Glu-B3. Due to resistance to some Lr26 virulent pathotypes, L224-3 may carry another resistance gene. Based on the genetic analysis using the pathotype FHBQ with virulence to Lr26 the resistance of L224-3 was controlled by a single dominant gene, tentatively designated LrL224. Four SSR markers (barc8, gwm582, wmc419, and wmc694) and one STS marker (ω-secali/Glu-B3) on 1B were closely linked to LrL224. The two flanking SSR loci were barc8 and gwm582, with the genetic distances of 4.3 and 4.6 cM, respectively. LrL224 was located on 1BL, and it showed different seedling reactions with other genes on 1B. Therefore LrL224 is likely to be a new leaf rust resistance gene.